title: RBE4 cells are highly resistant to paraquat-induced cytotoxicity: studies on uptake and efflux mechanisms.
abstract: Paraquat (PQ) is a widely used, highly toxic and non-selective contact herbicide, which has been associated with central neurotoxic effects, namely the development of Parkinson's disease, but whose effects to the blood-brain barrier (BBB) itself have rarely been studied. This work studied the mechanisms of PQ uptake and efflux in a rat's BBB cell model, the RBE4 cells. PQ is believed to enter cells using the basic or neutral amino acid or polyamine transport systems or through the choline-uptake system. In contrast, PQ efflux from cells is reported to be mediated by P-glycoprotein. Therefore, we evaluated PQ-induced cytotoxicity and the effect of some substrates/blockers of these transporters (such as arginine, L-valine, putrescine, hemicholinium-3 and GF120918) on such cytotoxicity. RBE4 cells were shown to be extremely resistant to PQ after 24 h of exposure; even at concentrations as high as 50 mM approximately 45% of the cells remained viable. Prolonging exposure until 48 h elicited significant cytotoxicity only for PQ concentrations above 5 mM. Although hemicholinium-3, a choline-uptake system inhibitor, significantly protected cells against PQ-induced toxicity, none of the effects were observed for arginine, L-valine or putrescine. Meanwhile, inhibiting the efflux pump P-glycoprotein using GF120918 significantly enhanced PQ-induced cytotoxicity. In conclusion, PQ used the choline-uptake system, instead of the transporters for the basic or neutral amino acids or for the polyamines, to enter RBE4 cells. P-glycoprotein extrudes PQ back to the extracellular medium. However, this efflux mechanism only partially explains the observed RBE4 resistance to PQ.
authors.affiliation: REQUIMTE, Laboratório de Toxicologia, Departamento de Ciências Biológicas, Faculdade de Farmácia, Universidade do Porto, Rua de Jorge Viterbo Ferreira, 228, Edifício 3, 4º piso, 4050-313, Porto, Portugal.
authors.forename: V
authors.initials: V
authors.lastname: Vilas-Boas
authors.forename: R
authors.initials: R
authors.lastname: Silva
authors.forename: P
authors.initials: P
authors.lastname: Guedes-de-Pinho
authors.forename: F
authors.initials: F
authors.lastname: Carvalho
authors.forename: M L
authors.initials: ML
authors.lastname: Bastos
authors.forename: F
authors.initials: F
authors.lastname: Remião
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D001812:Blood-Brain Barrier / Q000166:cytology / Q000187:drug effects; D002460:Cell Line; D002794:Choline / Q000378:metabolism; D042783:Endothelial Cells / Q000187:drug effects; D006540:Herbicides / Q000633:toxicity*; D010269:Paraquat / Q000493:pharmacokinetics / Q000633:toxicity*; D051381:Rats
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D006540:Herbicides; D002794:Choline; D010269:Paraquat
keywords: P-glycoprotein; RBE4 cells; blood-brain barrier; choline-uptake system; paraquat